Growth of cells on a perfluorocarbon-medium interphase: A quantitative assay for anchorage-independent cell growth

Summary A high density, purified, nontoxic solvent, heptacosafluorotributylamine (FC43), was successfully used as a culture surface for growing several normal and oncogene-transformed cell lines under anchorage-independent conditions. Normal rat kidney (NRK) fibroblasts and the normal mammary epithelial cell lines NMuMG and A1, clone N4, of murine and human origin, respectively, failed to grow at a FC43 growth medium interphase or in soft agar in the absence of transforming growth factor alpha (TGFα) and transforming growth factor beta (TGFβ). However, NRK fibroblasts transformed with the Kirstenras viral oncogene (K-NRK) or NMuMG cells transformed with a point-mutated c-Harvey-ras proto-oncogene or polyoma middle T-transforming gene (NMuMG-ras ^H and NMuMG-pyt, respectively) exhibited rapid growth and formed large colonies when cultured on an FC43-medium interphase. In addition, NRK cells treated with TGFα and TGFβ and K-NRK cells grown on FC43 exhibited a sensitivity to the growth inhibitory effects of 4-cis-L-hydroxyproline comparable to that observed for the same cells grown in soft agar. These results demonstrated that the two-phase assay system (FC43-growth medium interphase) may be superior to soft agar for monitoring the anchorage-independent growth of cells because of the ease of cell plating, the ability to recover cells and secreted products from the upper aqueous phase, and the shorter growth period required to complete the assay (3–4 days). Editor's Statement This report describes the application of a novel technique that provides an alternative approach to assay of anchorage independence with some unique advantages over conventional methods.